Penicillin acylases are enzymes which are produced by many different microorganisms including both bacteria and fungi as disclosed by Claridge et al., "Bacterial Penicillin Amidase." Nature 187, 237-238 (1960), Rolinson et al., "Formation of 6-Aminopenicillanic Acid from Penicillin by Enzymatic Hydrolysis." Nature 187, 236-237 (1960), and Sakaguchi et al., "A Preliminary Report on a New Enzyme Penicillin Amidase," J. Agr. Chem. Soc. Japan, 23, 411 (1950). These enzymes catalyze the hydrolysis of the phenylacetic acid side chain from benzyl penicillin to give 6-aminopenicillanic acid which is used commercially in the production of semisynthetic penicillins.
Bacillus strains, such as Bacillus megaterium ATCC 14945 are highly productive for penicillin acylase. Derivatives of Bacillus megaterium ATCC 14945 have been obtained using conventional mutagenesis and screening techniques which produce much higher industrial penicillin acylase yields than does ATCC 14945. However, techniques for producing penicillin acylase in even higher yields are still highly sought after.
Recombinant DNA technology has been used with success by many researchers in increasing the production of products specified by a single gene through the insertion of that gene on multi-copy plasmids in genetically well understood host organisms such as E. Coli, S. cerevisiae or B. subtilis. However, until now, no one has applied recombinant DNA technology to Bacillus megaterium to further improve industrial penicillin acylase yields.